Several lysosomal storage disorders, such as CHS or HPS, display similarities in terms of their clinical manifestations and immunologic mechanisms, one of them being the decreased or absent cytotoxic activity of NK cells resulting in immune deficiency. Thus, we are investigating whether impaired NK cell function in those disorders is associated with defects in secretory lysosome (lytic granule) morphology, biogenesis, motility, exocytosis, or combination of these factors. We analyzed NK cells from atypical CHS patients with missense mutations in the LYST ARM/HEAT or BEACH domains. CHS NK cells displayed severely reduced cytotoxicity. Mutations in the ARM/HEAT domain led to a reduced number of perforin-containing granules, which were significantly increased in size, able to polarize to the immunological synapse (IS), but unable to properly fuse with the plasma membrane. Mutations in the LYST BEACH domain resulted in the formation of normal or slightly enlarged granules that had markedly impaired polarization to the immunological synapse, but could be exocytosed upon reaching the IS. Perforin-containing granules in CHS NK cells did not acquire certain lysosomal markers (LAMP1/2), but were positive for markers of transport vesicles (CI-MPR), late endosomes (Rab27a), and to some extent, early endosomes (EEA-1), indicating impaired integrity of endo-lysosomal compartments. CHS NK cells had normal cytokine compartments and secretion, demonstrating that distinct domains of LYST are required for granule trafficking and exocytosis, but not constitutive secretion. Our data shows that LYST is involved in regulation of multiple aspects of NK cell lytic activity ranging from governance of lytic granule size to control of their polarization and exocytosis, as well as the regulation of endo-lysosomal compartment identity. Furthermore, we analyzed NK cells from patients diagnosed with HPS-1, HPS-2, HPS-4, and a newly identified HPS-10. NK cells from HPS-2 and HPS-10 patients had markedly reduced cytotoxic capabilities that correlated with defective NK cell degranulation, while NK cells from HPS type 1 and HPS type 4 patients had only slightly decreased capacity to kill the target cells. Microscopic analysis revealed that NK cells from patients with HPS-2 and HPS-10 had enlarged, granzyme A- and perforin-positive lytic granules that failed to translocate to the immunological synapse in response to target cell recognition; this translated to severely reduced granule exocytosis and deficient NK cell cytotoxicity. While both HPS-2 and HPS-10 NK cells shared a similar cellular phenotype in regard to the impairment of regulated exocytosis, they differed in the regulation of the constitutive secretion pathway. HPS-10 NK cells secreted normal amounts of TNF&#61537; and IFN&#61543;, whereas HPS-2 NK cells displayed defective cytokine exocytosis, and accumulated cytokines inside of the cell. Thus, HPS-10 affects the cytolytic function of NK cells, while HPS-2 affects both lytic granule and cytokine secretion.